Reactions were performed with an initial step of AmpliTaq Platinum activation at 95C for 10 min and 40 cycles (melting phase 94C for 40 sec, annealing 60C for 30 sec and extension 68C for 45 sec). NONOate (DETA/NO), resulted in a lower capacity of the cells to synthesize cGMP in response to a subsequent NO stimulus. This effect was not prevented by an increase of intracellular reduced glutathione level. DETA/NO treatment decreased sGC subunit mRNA and 1 subunit protein levels. Both sGC activity and 1 subunit levels decreased more rapidly in chromaffin cells exposed to NO than in cells exposed to the protein synthesis inhibitor, cycloheximide, suggesting that NO decreases 1 subunit stability. The presence of cGMP-dependent protein kinase (PKG) inhibitors efficiently prevented the DETA/NO-induced down rules of sGC subunit mRNA and partially inhibited the reduction in 1 subunits. Conclusions These results suggest that activation of PKG mediates the drop in sGC subunit mRNA levels, and that NO down-regulates sGC activity by reducing subunit mRNA levels through a cGMP-dependent mechanism, and by reducing 1 subunit stability. Background The soluble Carmustine form of guanylyl cyclase (sGC) is the main receptor for the signaling agent nitric oxide (NO) [1,2]. This signaling molecule performs important physiological and biochemical functions in the cardiovascular, pulmonary and neural systems, activating sGC and thus increasing cGMP levels [3-6]. In certain conditions and disease claims such as hypoxia and hypertension, a disturbance in sGC levels and/or activity may play a crucial part in the pathophysiology of these disorders [7-9]. Moreover, desensitization of sGC may also be involved in tolerance to NO when this compound is used for restorative purposes [10,11]. Purification of mammalian sGC yields a heterodimer comprised of two subunits and of which four types exist (1, 2, 1, 2) [12-17]. Structurally, each subunit has a C-terminal cyclase catalytic website, a central dimerization region and a N-terminal portion. This last portion constitutes the heme-binding website and represents the least conserved region of the protein. Cloning and manifestation experiments have shown that both and subunits are required for sGC to be functionally active [18,19]. In mammalian cells, two different heterodimeric enzymes, 1/1 and 2/1, have been detected, and although 1 and 2 differ in their main structure, the two heterodimers were found to be functionally alike [18,20]. Recently, it has been shown the activation of 2 subunit of sGC by NO and this enzyme has a monomeric structure [21]. While much has been learned about the rules of NO synthase [22,23], there is scare data on sGC rules, despite its essential part in the actions mediated by endogenous or exogenous NO [17,24]. Different reports have shown a decrease in sGC activity after pre-treatment of cells or cells with NO-releasing compounds, or a higher sensitivity of the enzyme when endogenous NO synthesis is definitely GTF2F2 inhibited [10,25,26]. However, the mechanism involved in this phenomenon remains unclear. Redox reactions may be a plausible mechanism for enzyme desensitization, as several studies have shown the redox state of the enzyme bound heme and protein thiol groups has a major part in controlling enzyme activity [10,27,28]. This activity can also be controlled by a phosphorylation/dephosphorylation mechanism [29,30], and there is growing evidence that sGC activity is definitely controlled both in the protein and mRNA levels. Several authors have reported that treatments, such as forskolin, dibutyryl-cAMP, 3-isobutyl-methyl xanthine Carmustine [31,32], endotoxin and/or IL-1 [33,34], NO donating compounds [11,26] or nerve growth element [35] affect sGC mRNA levels in various cell types. The NO/cGMP pathway has been established as a major controller of several physiological functions of the nervous system [6,36]. Moreover, the effects of NO/cGMP on neuronal differentiation and survival, and synaptic plasticity suggest that this transmission transduction pathway regulates gene manifestation in the nervous system [37]. Since the part Carmustine of sGC in transducing inter- and intracellular signals conveyed by NO is definitely pivotal, knowledge of the molecular mechanisms involved in sGC rules may help our understanding of the physiological and pathophysiological significance of this transmission transduction pathway in the nervous system. Compared with findings in vascular cells, little is known about the effect of prolonged exposure of neural cells to nitric oxide on subsequent NO stimulation of these cells, and the capacity of this agent to elicit cGMP raises. The aim of the present study was to establish whether long term treatment of chromaffin cells with low doses of nitric oxide affects sGC activity inside a widely used bovine neural cell model. The findings presented suggest that chronic exposure to NO decreases sGC activity by reducing the availability of 1 subunit.